Study On The Effect And Mechanism Of Deep-fried Atractylodis Rhizoma On Anti-diarrhea Based On Intestinal Flora

1 PURPOSEDeep-fried Atractylodis Rhizoma is a more commonly used spleen-invigorating and antidiarrheal drug,mainly used for embolism of chronic diarrhea and chronic dysentery caused by spleen deficiency.At present,there are few studies on the role and mechanism of deep-fried Atractylodis Rhizoma antidiarrheal effect.In this study,by comparing the anti-diarrhea effect of different extracts of deep-fried Atractylodis Rhizoma,the main anti-diarrhea effective parts of deep-fried Atractylodis Rhizoma were clarified,and the molecular mechanism of anti-diarrhea effect of deep-fried Atractylodis Rhizoma was further studied to preliminary clarify its mechanism of drug effect,so as to provide reference for further expanding the clinical application of deep-fried Atractylodis Rhizoma.2 METHODS2.1 Study on the screening of anti diarrhea effect site of deep-fried Atractylodis RhizomaWistar rats were used as research objects,and the model of spleen deficiency diarrhea was made by rhubarb.The high and low doses of alcohol extract of deep-fried Atractylodis Rhizoma,water extract of deep-fried Atractylodis Rhizoma and Shen-Ling-Bai-zhu-San were given respectively.The body weight,fecal water content,diarrhea grade score were used as indexes.The colonic pathological sections were observed and the anti-diarrhea effects of each group were investigated.2.2 Study on anti-diarrhea effects of deep-fried atractylodis rhizomaFirst,Wistar rats were divided into 5 groups,10 in each group,which were normal group,model group,high and low-dose group of alcohol extract of deep-fried Atractylodis Rhizoma,Shen-Ling-Bai-zhu-San group.Except the normal group,all the other groups were modeled as diarrhea by rhubarb,and then continued treatment for 7 days.with body weight,fecal water content,diarrhea grade score,motilin MTL,gastrin GAS,serum tumor necrosis factor TNF-α,interleukin IL-10 were used as indicators.The pathological sections of intestinal acupoints were used to further investigate the anti-diarrhea effect of alcohol extract of deep-fried Atractylodis Rhizoma.2.3 Study on the anti diarrhea mechanism of alcohol extract of deep-fried atractylodis rhizoma based on intestinal permeabilityWistar rats were divided into 5 groups,10 rats in each group,which were normal group,model group,high and low-dose group of alcohol extract of deep-fried Atractylodis Rhizoma,Shen-Ling-Bai-zhu-San group.Except the normal group,the other groups all used rhubarb to make spleen deficiency diarrhea model and then continued treatment for 7 days.After the experiment,serum was taken to measure 5-HT,colonic tissue was taken to measure the expression of 5-HT3 R,AQP3 and AQP8.The expression of connexin ZO-1and occludin,myosin light chain kinase and myosin light chain,and the p38 MAPK signal pathway were detected to further clarify the anti diarrhea mechanism of alcohol extract of deep-fried atractylodis rhizoma.2.4 Study on the effect of alcohol extract of deep-fried atractylodis rhizoma on intestinal flora based on 16 Sr DNAWistar rats were divided into 5 groups,10 rats in each group,which were normal group,model group,high and low-dose group of alcohol extract of deep-fried Atractylodis Rhizoma,Shen-Ling-Bai-zhu-San group.Except the normal group,the other groups all used rhubarb to make spleen deficiency diarrhea model and then continued treatment for 7 days.After the experiment,the colon contents or feces of rats were taken with sterilized EP tube and stored in-80℃or liquid nitrogen.16 s RNA technology was used to analyze the microorganism in the intestinal contents or feces of rats,to clarify its structure and abundance composition,and to further clarify the effect of deep-fried Atractylodis Rhizoma on the intestinal microecology.3 RESULTS3.1 Study on the screening of anti diarrhea effect site of deep-fried Atractylodis RhizomaCompared with the model group,the body weight of deep-fried Atractylodis Rhizoma water extract and deep-fried Atractylodis Rhizoma alcohol extract high-dose group increased significantly.In terms of fecal water content and diarrhea grade scores,both of them can reduce fecal water content and diarrhea grade scores,but the effect of deep-fried Atractylodis Rhizoma alcohol extract high-dose group is significantly better than that of water extract high-dose group.In the aspect of reducing intestinal inflammation,the high-dose alcohol extract group was the best,followed by the high-dose water extract group,while the other groups had no significant difference.In conclusion,under the same dose,the anti diarrhea effect of alcohol extract of deep-fried Atractylodis Rhizoma is significantly better than that of water extract,so it is preliminarily determined that the main anti diarrhea part of deep-fried Atractylodis Rhizoma is its alcohol extract part.3.2 Study on the effect of alcohol extract of deep-fried Atractylodis Rhizoma on anti diarrheaCompared with the model group,the body weight of the high-dose alcohol extract group of deep-fried Atractylodis Rhizoma increased significantly,and the fecal water content and diarrhea grade scores decreased significantly.In addition,it was found that ethanol extract of deep-fried Atractylodis Rhizoma can not only reduce intestinal inflammation and protect intestinal injury,but also reduce the overall inflammatory level in rats.In the aspect of gastrointestinal motility hormone,deep-fried Atractylodis Rhizoma can also increase the expression of gastrointestinal motility hormone and improve gastrointestinal dysfunction.Through the further study on the antidiarrheal effect of deep-fried Atractylodis Rhizoma,we found that deep-fried Atractylodis Rhizoma can not only significantly alleviate diarrhea caused by spleen deficiency,but also inhibit the occurrence of various kinds of inflammation in the body,and also recover the gastrointestinal dysfunction caused by spleen deficiency.3.3 Study on the anti diarrhea mechanism of alcohol extract of deep-fried atractylodis rhizoma based on intestinal permeabilityIt was found that the alcohol extract of deep-fried Atractylodis Rhizoma could reduce the expression of 5-HT and 5-HT3 R receptor in rats.In addition,the high dose group of deep-fried Atractylodis Rhizoma could increase the expression of AQP3 and AQP8,and recover the abnormal water metabolism.In terms of protecting intestinal injury,it was found that the high dose group of deep-fried Atractylodis Rhizoma can not only promote the integrity of intestinal barrier,increase the expression of tight junction protein ZO-1 and occludin,but also inhibit the phosphorylation of MLCK and MLC,reduce the expression level of PAR-2,thus promote the expression of tight junction protein and further protect the intestinal tract.3.4 Study on the effect of alcohol extract of deep-fried atractylodis rhizoma on intestinal flora based on 16 Sr DNAThrough the study of intestinal microecology in each group of rats,it was found that the composition of intestinal microflora in each group was different,but Bacteroidetes and Firmicutes were the main composition colonies.In the model group,the abundance of some genera,such as Clostridium,Parabacteroides and Helicobacter pylori,were significantly increased in the rats Bacteria can cause diarrhea.After the treatment of deep-fried Atractylodis Rhizoma,these harmful bacteria all decreased in varying degrees.In the high-dose group,the species richness of the beneficial flora such as akkermansia and lactobacilli increased significantly,which indicated that the ethanol extract could promote the formation of beneficial flora and alleviate diarrhea by inhibiting the production of harmful bacteria.4 CONCLUSIONFirst of all,through the comparative study of different extracts of deep-fried Atractylodis Rhizoma,it is preliminarily confirmed that the main antidiarrheal part of deep-fried Atractylodis Rhizoma is alcohol extract.Then through a more comprehensive index evaluation of the antidiarrheal efficacy of the alcohol extract of deep-fried Atractylodis Rhizoma and its mechanism were explored.The results showed that the alcohol extract of deep-fried Atractylodis Rhizoma can not only effectively alleviate diarrhea caused by spleen deficiency,but also improve intestinal pathological changes,inhibit the decrease of tight junction protein and protect the integrity of intestinal barrier through P38 MAPK signal pathway.In addition,it was also found that the ethanol extract of deep-fried Atractylodis Rhizoma could regulate the intestinal flora,improve the intestinal flora structure and restore the intestinal microecology.The above results are helpful for us to understand the therapeutic effect and mechanism of deep-fried Atractylodis Rhizoma,and provide important basis for the rational application of deep-fried Atractylodis Rhizoma.